This is a competing renewal of an R01 grant to determine how gamma-herpesvirus replication is controlled, both during reactivation from latency as well as following virus entry into a permissive cell. Gamma-herpesviruses are associated with the development of lymphoproliferative disorders and lymphoma, particularly in immunosuppressed individuals. A detailed understanding of how virus replication is triggered is critical to understanding the biology of gamma-herpesvirus infections, and may identify possible targets for interfering with viral persistence in the host. In this renewal application we propose to investigate the regulation of murine gamma-herpesvirus; 68 (MHV68) replication. MHV68 infection of mice provides a tractable small animal model system for characterizing the role of specific genes in viral pathogenesis and maintenance of chronic infection. Studies on control of MHV68 replication will focus on the following aims: Aim 1: Regulation of gene 50 expression Aim 1.a. Identify critical cis-elements and trans-acting factors regulating gene 50 transcription; and. Aim 1.b. Determine role of proximal and distal gene 50 promoters in MHV68 reactivation and replication. Aim 2: Regulation of Rta function and early gene expression Aim 2.a. Determine role of orf49 gene product in Rta activation of target promoters in B cells and non-B cells; Aim 2.b. Identify immediate-early MHV68 genes involved in virus reactivation from B cells; and Aim 2.c. Characterize impact of BLIMP-1 and XBP-1 on viral gene expression in B cells. Aim 3: Role of mLANA in virus reactivation and replication. Aim 3.a. Identify functional domains in mLANA that contribute to virus reactivation/replication; Aim 3.b. Identify and characterize mLANA associated proteins; and Aim 3.c. Assess whether functional domains in KSHV LANA and mLANA are interchangable.